This project aims to study the conformational changes in the contractile proteins of the thin filaments -- actin, tropomyosin, and the troponin complex and subunits --- using fluorescent probes with long lifetimes of the excited state. Initially proteins will be prepared from rabbit muscle, but later proteins from dog hearts will be used. The changes associated with reconstitution of the thin filament proteins, with interaction with Ca ions, and with heavy meromyosin and ATP will be studied. Comparisons will be made with proteins from failing hearts and cultured embryonic hearts will be used for drug studies. We hope to quantify the structural changes in the contractile proteins which take place upon association with the hope of understanding better the nature of the myosin-thin filament interaction, the mechanism of activation by Ca ions of the active contractile state, and what changes, if any, are found in pathological muscle and the effects of drugs. The use of time-resolved fluorescence spectrometry is central to the performance of these experiments. The methodologies of data analysis for deconvolution and analysis of multi-exponential curves are being improved, and a computer simulation of Brownian motion for flexible molecules is being developed to obtain hydrodynamic data from time-dependent anisotropy of fluorescence polarization by numerical analysis.